The primate lentiviruses, including human immunodeficiency virus (HIV) type 1 (HIV-1), and type 2 (HIV-2) and SIV are genetically, structurally and functionally similar. HIV-1 and HIV-2 are genetically related, antigenically cross reactive, and share a common cellular receptor (CD4). See, Rosenburg and Fauci (1993) in Fundamental Immunology, Third Edition Paul (ed) Raven Press, Ltd., New York (Rosenburg and Fauci 1) and the references therein for an overview of HIV infection. Due to the pandemic spread of HIV-1 (and increasingly, HIV-2), an intense world-wide effort to unravel the molecular mechanisms and life cycle of these viruses is underway. It is now clear that the life cycle of these viruses provide many potential targets for inhibition by gene therapy, including cellular expression of transdominant mutant gag and env nucleic acids to interfere with virus entry, TAR (the binding site for tat, which is typically required for transactivation) decoys to inhibit transcription and trans activation, and RRE (the binding site Rev; i.e., Rev Response Element) decoys and transdominant Rev mutants to inhibit RNA processing. See, Wong-Staal et al., PCT/US94/05700; Rosenburg and Fauci (1993) in Fundamental Imnnunology, Third Edition Paul (ed) Raven Press, Ltd., New York and the references therein for an overview of HIV infection and the HIV life cycle, gene therapy vectors utilizing ribozymes, antisense molecules, decoy genes, transdominant genes and suicide genes, including retroviruses. See also, Yu et al., Gene Therapy (1994) 1:13-26. Antisense and ribozyme therapeutic agents are of increasing importance in the treatment and prevention of HIV infection.
Antisense gene therapeutic agents and ribozymes are entering clinical trials as gene therapeutic agents for the treatment of HIV infection. Ribozymes are particularly potent therapeutic agents because (i) as RNA molecules, they are not likely to induce host immunity that eliminates the transduced cells; (ii) although they resemble antisense molecules in their sequence specific recognition of target RNA, their ability to cleave the target RNA catalytically renders them more efficient than simple anti-sense molecules; and (iii) they can potentially cleave both afferent and efferent viral RNA, and therefore inhibit both preintegration and postintegration steps of the virus replication cycle. T-cell lines (Yamada et al., Gene Therapy (1994) 1:38-45) and primary lymphocytes (Leavitt et al., Hum. Gene Ther. (1994) 5:1115-1120) transduced with retroviral vectors expressing anti-HIV hairpin ribozyrnes are resistant to exogenous infection with diverse strains of HIV-1. Furthermore, macrophages derived from primary CD34.sup.+ hematopoietic stem/progenitor cells were also resistant to challenge with a macrophage tropic strain of HIV-1 (Yu et al., Virology (1995) 206:381-386).
Because of the dramatic potential of gene therapy, constructs and methods which improve the efficacy of viral inhibitors used in gene therapy are of increasing importance. The present invention provides methods and compositions which are optionally combined with other viral inhibitors, compounds and methods to provide cells with enhanced viral resistance. The present invention also provides diagnostic reagents and methods, and kits based upon the compositions and methods of the invention.